Roles of Noncoding RNAs in Regulation of Mitochondrial Electron Transport Chain and Oxidative Phosphorylation

The mitochondrial electron transport chain (ETC) plays an essential role in energy production by inducing oxidative phosphorylation (OXPHOS) to drive numerous biochemical processes in eukaryotic cells. Disorders of ETC and OXPHOS systems are associated with mitochondria- and metabolism-related diseases, including cancers; thus, a comprehensive understanding of the regulatory mechanisms of ETC and OXPHOS systems is required. Recent studies have indicated that noncoding RNAs (ncRNAs) play key roles in mitochondrial functions; in particular, some ncRNAs have been shown to modulate ETC and OXPHOS systems. In this review, we introduce the emerging roles of ncRNAs, including microRNAs (miRNAs), transfer-RNA-derived fragments (tRFs), long ncRNAs (lncRNAs), and circular RNAs (circRNAs), in the mitochondrial ETC and OXPHOS regulation.


Introduction
Mitochondria play a crucial role in a diverse range of biological processes in eukaryotic cells. One of the important functions of mitochondria is to generate energy in the cell via electron transport and oxidative phosphorylation (OXPHOS). The mitochondrial electron transport chain (ETC) consists of a series of electron carriers embedded in the inner mitochondrial membrane. Electrons from NADH and FADH 2 , which are produced in the metabolic pathways of glucose and other nutrients, are transferred along the ETC, leading to the transport of protons across the membrane from the mitochondrial matrix to the intermembrane space. This establishes a proton gradient across the inner mitochondrial membrane, which is then used by the ATP synthase enzyme to produce ATP via OXPHOS. OXPHOS is the process of phosphorylating ADP to form ATP using energy from the proton gradient established by the ETC. The ETC comprises a series of four enzymatic complexes that contain complex I (NADH ubiquinone oxidoreductase/NADH dehydrogenase), complex II (succinate ubiquinone oxidoreductase/succinate dehydrogenase), complex III (ubiquinol cytochrome c oxidoreductase/cytochrome bc1 complex), and complex IV (cytochrome c oxidase). OXPHOS is accomplished by complex V (ATP synthase) [1]. Some of these complexes generate higher-order assemblies called 'supercomplexes' [1,2] (Figure 1). In mammalian cells, a supercomplex called 'respirasome' is the major structure, which is composed of one complex I, two complex IIIs, and one complex IV (I + III 2 + IV). Although the biological function and regulation of the mitochondrial supercomplex assembly is highly debated, it can contribute to efficient energy production [3][4][5][6], reducing reactive oxygen species (ROS) [7][8][9], and the stability of complex I [10]. reactive oxygen species (ROS) [7][8][9], and the stability of complex I [10]. The ETC complexes and complex V are composed of specific protein subunits that are encoded in nuclear DNA (nDNA) or mitochondrial DNA (mtDNA) [11]. Some mutations of these subunits as well as assembly factors of ETC complexes and complex V are known to cause the dysregulation of mitochondrial functions associated with mitochondrial diseases such as mitochondrial encephalopathy, resulting in lactic acidosis and stroke-like episodes (ME-LAS), and Leigh syndrome. Furthermore, ETC/OXPHOS disorders are assumed to result in metabolism-related diseases and cancers [12][13][14][15][16]. Thus, a comprehensive understanding of ETC/OXPHOS regulation is crucial for precise diagnosis of and therapeutic intervention in mitochondria-and metabolism-related diseases and cancers.

Figure 1.
Mitochondrial electron transport chain complexes, supercomplex, and OXPHOS. The mitochondrial electron transport chain (ETC) is operated by four enzymatic protein complexes in the inner mitochondrial membrane, i.e., complex I (NADH ubiquinone oxidoreductase/NADH dehydrogenase; CI), complex II (succinate ubiquinone oxidoreductase/succinate dehydrogenase; CII), complex III (ubiquinol cytochrome c oxidoreductase/cytochrome bc1 complex; CIII), and complex IV (cytochrome c oxidase; CIV). Complexes I-IV mediate electron transport in the sequence from NADH or FADH2 to molecular oxygen. Through the ETC, protons (H + ) are transported across the inner mitochondrial membrane to the intermembrane space. The established proton gradient is used to generate ATP from ADP phosphorylation by complex V (ATP synthase; CV), which is known as electron transport-linked phosphorylation or oxidative phosphorylation (OXPHOS). A certain portion of the mitochondrial electron transport complexes form higher-order structures called 'supercomplexes'. The supercomplex generated by one CI, two CIIIs, and one CIV has been termed the 'respirasome'. Mitochondrial electron transport chain complexes, supercomplex, and OXPHOS. The mitochondrial electron transport chain (ETC) is operated by four enzymatic protein complexes in the inner mitochondrial membrane, i.e., complex I (NADH ubiquinone oxidoreductase/NADH dehydrogenase; CI), complex II (succinate ubiquinone oxidoreductase/succinate dehydrogenase; CII), complex III (ubiquinol cytochrome c oxidoreductase/cytochrome bc 1 complex; CIII), and complex IV (cytochrome c oxidase; CIV). Complexes I-IV mediate electron transport in the sequence from NADH or FADH 2 to molecular oxygen. Through the ETC, protons (H + ) are transported across the inner mitochondrial membrane to the intermembrane space. The established proton gradient is used to generate ATP from ADP phosphorylation by complex V (ATP synthase; CV), which is known as electron transport-linked phosphorylation or oxidative phosphorylation (OXPHOS). A certain portion of the mitochondrial electron transport complexes form higher-order structures called 'supercomplexes'. The supercomplex generated by one CI, two CIIIs, and one CIV has been termed the 'respirasome'.
ing the translation of cytochrome c oxidase subunit 1 (COX1) and reducing complex IV's function [17]. In another type of ncRNA, namely, long ncRNAs (lncRNAs), metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been suggested to promote hepatocellular carcinoma (HCC) cell proliferation, migration, and invasion by epigenetically regulating mtDNA-encoded ETC-related genes [18]. These ncRNAs may be promising targets in the diagnosis and treatment of related diseases [19,20].
In this review, we summarize the functional roles of ncRNAs, including miRNAs, transfer RNA-derived fragments (tRFs), lncRNAs, and circular RNAs (circRNAs), in the cross-talk signaling between mitochondria and the nucleus targeting regulated ETC complexes in mammalian cells.

Species and Classification of ncRNAs Affecting Mitochondria-Nucleus Crosstalk
In mammalian cells, numerous ncRNAs are synthesized to exhibit their various functions, including RNA splicing, RNA modification, and mRNA translation. ncRNAs are classified according to their length and structures. Small ncRNAs (sncRNAs) are classified as ncRNAs that are shorter than 200 nucleotides. miRNAs are a class of sncRNAs that are approximately 22 nucleotides long. miRNAs are synthesized as primary transcripts in the nucleus and are subsequently cleaved by the DROSHA/DiGeorge syndrome critical region 8 (DGCR8) complex to form single stem-loop precursor molecules, which are known as pre-miRNAs. Pre-miRNAs are generally transported to the cytoplasm by a nuclear transport receptor, Exportin 5, along with a cofactor RanGTP, and are then further cleaved by DICER1 to generate mature double-stranded miRNA duplexes [21]. In most cases, the miRNA duplex is incorporated into the RNA-binding protein argonaute 2 (AGO2) in mammalian cells; then, only one RNA strand (the guide strand) forms a complex with AGO2, leading to the formation of the miRNA-induced silencing complex (miRISC). The miRISC typically recognizes the 3 untranslated region (3 UTR) of target mRNAs using the guide strand and downregulates the translation and stability of these mRNAs. [22][23][24][25]. Besides their role in negative post-transcriptional regulation, miRNAs have been proposed to possess non-canonical functions outside the cytoplasm. For example, some miRNAs may act as chromatin and transcriptional regulators in the nucleus and as translational activators in mitochondria [26,27]. Mitochondrially located miRNAs are commonly termed "mitomiRs", which are derived from either transcripts of mitochondria or the nuclear genome [28,29]. Nuclear genome-encoded mitomiRs are more abundant in mitochondria compared to mitochondrial genome-encoded genes. The nuclear genome-encoded mitomiRs are often located in loci within or close to mitochondrial genes so that their transcriptions can be coregulated.
tRNA derived fragments (tRFs) are another class of sncRNAs, which are approximately 16-35 nt in length and originate from the cleavage of precursor or mature tRNAs by specific nucleases under certain conditions including cellular stress [30][31][32][33][34][35][36][37]. tRFs can be derived from tRNAs found in both the nucleus and mitochondria [38][39][40][41][42][43]. In the human genome, more than 400 nuclear genes encode for approximately 280 different tRNA isotypes, and mitochondrial DNA encodes 22 tRNAs [44][45][46]. The diversity of tRNA molecules can be attributed to the presence of isoacceptors, which are tRNAs that carry the same amino acid but have different nucleotide sequences in both the body and the anticodon region, as well as isodecoders, which are tRNAs that possess the same anticodon and carry the same amino acid but differ in their nucleotide sequences in the body. Isoacceptors and isodecoders exist in several copies in the genome and their expressions are tissue-and cell-type-specific, which can be modulated by stress conditions. Post-transcriptional tRNA modifications can take place in the nucleus, cytosol, and mitochondria and are induced by various tRNA-modifying enzymes through modulating base pairing, tRNA folding, and stability [47]. On average, cytoplasmic and mitochondrial tRNAs in mammals contain 13 and 5 modified bases, respectively. Hypomodified tRNAs are targeted for degradation and undergo endonuclease-mediated fragmentation. tRFs resulting from the fragmentation of mature tRNAs are classified in six major structural categories based on their positions in the parental tRNA sequence, as follows: 5 -tRFs and 3 -tRFs generated from either distal end of mature tRNAs, internal tRFs (i-tRFs) from internal sequences of mature tRNAs, 5 -tRNA halves and 3 -tRNA halves from the cleavage of anticodon sites of mature tRNAs (tRNA-derived stress-induced RNA (tiRNA)-5s and -3s, respectively), and tRF-1s generated from precursor tRNAs. While angiogenin is the enzyme primarily responsible for cleaving the anticodon site, different endonucleases such as DICER, SLNF13, RNase T2, and RNase Z have been reported to generate other classes of tRFs [48][49][50]. tRF is often stimulated under stress conditions, in which post-transcriptional modifications of tRNA are impaired and function to repress gene expression in a manner similar to that of miRNAs [51,52]. Moreover, tRFs function as important modulators of many aspects of biological processes, including translation, apoptosis, immunity, nutritional uptake, and oxidative stress. Notably, with the innovative development of high-throughput sequencing techniques and bioinformatic analysis programs, it has become evident that nuclear genome-encoded tR-NAs and mitochondrial tRNAs (mt-tRNAs) can produce tRFs [43,[53][54][55]. mt-tRNA-derived tRFs (mt-tRFs) actively participate in intracellular communication and mitochondrial pathophysiology [55][56][57].
Noncoding RNAs longer than 200 nucleotides are referred to as lncRNAs. LncRNAs are transcribed from both nuclear and mitochondrial genomes, and there are approximately 20,000 lncRNA genes in humans, showing a diversity comparable to that of protein-coding genes. Although most lncRNAs remain functionally unannotated, some lncRNAs are thought to play essential roles in cell physiology and numerous biological processes and diseases, including various types of cancers [20,[58][59][60][61]. Some nuclear genome-encoded lncRNAs have been found in mitochondria. For example, one nuclear genome-encoded lncRNA, namely, RNA component of mitochondrial RNA processing endoribonuclease (RMRP), is imported to the mitochondrial matrix [62]. This lncRNA was identified as a component of the mitochondrial RNA-processing endoribonuclease (MRP) and has been suggested to be involved in the generation of primers for mitochondrial DNA replication [63]. Meanwhile, the role of RMRP in mitochondria is debated, as recent studies have suggested that RNA primer generation is mediated by the premature arrest of RNA polymerase mitochondrial (POLRMT), and the 3 half of this lncRNA (~130 nt) has been predominantly detected inside mitochondria [64,65].
CircRNAs are defined by their structure and not by their length. Such structures are single-stranded and covalently closed at the 5 and 3 ends [66,67]. The production of circRNA generally occurs through a distinct form of alternative splicing referred to as back-splicing. This process involves the 3 end of an exon joining to the 5 end of its own or a preceding exon through a 3 ,5 -phosphodiester bond, resulting in a circular structure with a back-splicing junction site [68][69][70]. The unique structure of circRNAs provides them with a longer-half life and greater resistance to exonucleases than linear RNAs [68,69,[71][72][73]. An increasing number of investigations uncovered diverse roles of circRNAs, such as acting as protein scaffolds or miRNA sponges, as well as having the ability to undergo translation into polypeptides [69,74,75]. Recently, mitochondria-located circRNAs have been discovered, and their functions have been investigated, which has expanded our understanding of circRNA derivation and mitochondrial transcriptome [76][77][78].
Regarding the targeting and import of nDNA-encoded ncRNAs into mammalian mitochondria, some proteins have been indicated as being involved in these processes. For example, polynucleotide phosphorylase (PNPASE) has been suggested to modulate the import of some ncRNAs, including RNase P RNA and RMRP. However, the mechanisms whereby nDNA-encoded ncRNAs target and are imported into mitochondria remain controversial, and thus new techniques are needed to precisely analyze nuclear-mitochondrial RNA dynamics (reviewed in [64]).
In this review, we define mitochondrial ncRNAs (mt-ncRNAs) as ncRNAs that are either transcribed from mtDNA regardless of subsequent mitochondrial localization or are mitochondria-located ncRNAs transcribed from nDNA. Another classification for ncRNAs that are neither located in mitochondria nor encoded by the mitochondrial genome but regulate mitochondrial functions is "mitochondria-associated ncRNAs". Mitochondriaassociated ncRNAs, a large portion of which are miRNAs, regulate mitochondrial functions by targeting nuclear genome-encoded mitochondrial mRNAs in the cytoplasm (Table 1, Figure 2). It has been suggested that mt-ncRNAs and mitochondria-associated noncoding RNAs regulate mitochondria-nucleus crosstalk through anterograde and retrograde signals [79,80]. . Noncoding RNAs regulating mitochondria-nucleus crosstalk. According to their localization and genome origin, mitochondrial noncoding RNAs (mt-ncRNAs) can be classified into four types: (a) nuclear-genome-encoded mitochondria-located mt-ncRNAs, (b) mtDNA-encoded mitochondria-located mt-ncRNAs, (c) mtDNA-encoded nucleus/cytosol-located mt-ncRNAs, and (d) nuclear-genome-encoded nucleus/cytosol-located mt-ncRNAs (termed "mitochondria-associated noncoding RNAs"). mt-ncRNAs regulate the mitochondrial electron transport chain and OXPHOS, which coordinate the crosstalk between nucleus and mitochondria as well as mitochondria-associated noncoding RNAs.

Roles of microRNAs in the Regulation of Mitochondrial ETC Complexes and OXPHOS
Numerous mitomiRs have been shown to affect the levels of enzymes encoded by the mitochondrial genome in ETC complexes (Table 2 and Figure 3). For instance, the mitochondria-located miRNA let-7a in MCF-7 breast cancer cells decreases the activity of ETC complex I by inducing the destabilization of the mRNA encoding NADH ubiquinone oxidoreductase core subunit 4 (ND4), which is a subunit of complex I [81].  Noncoding RNAs regulating mitochondria-nucleus crosstalk. According to their localization and genome origin, mitochondrial noncoding RNAs (mt-ncRNAs) can be classified into four types: (a) nuclear-genome-encoded mitochondria-located mt-ncRNAs, (b) mtDNA-encoded mitochondria-located mt-ncRNAs, (c) mtDNA-encoded nucleus/cytosol-located mt-ncRNAs, and (d) nuclear-genome-encoded nucleus/cytosol-located mt-ncRNAs (termed "mitochondria-associated noncoding RNAs"). mt-ncRNAs regulate the mitochondrial electron transport chain and OXPHOS, which coordinate the crosstalk between nucleus and mitochondria as well as mitochondria-associated noncoding RNAs.

Roles of microRNAs in the Regulation of Mitochondrial ETC Complexes and OXPHOS
Numerous mitomiRs have been shown to affect the levels of enzymes encoded by the mitochondrial genome in ETC complexes (Table 2 and Figure 3). For instance, the mitochondria-located miRNA let-7a in MCF-7 breast cancer cells decreases the activity of ETC complex I by inducing the destabilization of the mRNA encoding NADH ubiquinone oxidoreductase core subunit 4 (ND4), which is a subunit of complex I [81].  tiRNA-5 His(GTG) Rat Nuclear Decreases MPC1 and is involved in proliferation and insulin secretion of neonatal rat pancreatic β cells [93] tiRNA-5 Glu(CTC) Rat Nuclear Decreases MPC1 and is involved in proliferation and insulin secretion of neonatal rat pancreatic β cells [93] Mitochondria-associated tRFs tiRNA-5 Glu(TTC) Rat Nuclear Decreases MPC1 levels and is involved in mitochondrial respiration, proliferation, and insulin secretion of neonatal rat pancreatic β cells [93] lncRNA mt-lncRNAs

MALAT1
Human Nuclear Modulates the CpG methylation of mtDNA and the expression of mtDNA-encoded ETC-related genes in HepG2 HCC cells [18] SAMMSON Human Nuclear Modulates the expression of mtDNA-encoded ETC complex subunits by facilitating mitochondrial localization of p32 in melanoma cells [94,95] SAMMSON silencing increases the expression of ETC complexes, cellular respiration, and mitochondrial replication in doxorubicin-resistant MCF-7 breast cancer cells [96]  It has been suggested that miR-762 translocates into mitochondria in murine cardiomyocytes upon anoxia/reoxygenation treatment and decreases complex I activity and intracellular ATP levels, increases ROS levels, and enhances apoptotic cell death through suppressing the expression of NADH ubiquinone oxidoreductase chain 2 (ND2), which is a subunit of complex I [82]. Intriguingly, the administration of an inhibitor (antagomir) of miR-762 ameliorated myocardial ischemia/reperfusion injury in mice. Another example of the negative regulation of ETC complexes is provided by miR-181c, which binds to the 3 UTR of cytochrome c oxidase subunit 1 (COX1) mRNA, encoding a subunit of complex IV and suppressing COX1 translation in rat cardiac myocytes. The overexpression of miR-181c dramatically decreased COX1 protein levels, leading to complex IV remodeling and altering mitochondrial function, including by precipitating augmented ROS production [17]. Notably, the systemic administration of miR-181c in rats diminished exercise capacity and caused heart failure [101]. For patients with heart failure with a preserved ejection fraction (HFpEF), exercise training is a treatment that commonly used to improve such patients' aerobic capacity and quality of life. A recent study has shown that the circulating level of miR-181c is significantly higher in low responders to exercise training, suggesting that miR-181c may affect the exercise adaptation process in HFpEF patients partly through ETC regulation [102].  Table 2 in mitochondrial ETC and OXPHOS are illustrated herein. miRNAs, tRFs, lncRNAs, and circRNAs are represented in blue, red, green, and orange, respectively. Ndufa1, NADH ubiquinone oxidoreductase subunit A1; Ndufa2, NADH ubiquinone oxidoreductase subunit A2; Ndufa3, NADH ubiquinone oxidoreductase subunit A3; Ndufab1, NADH ubiquinone oxidoreductase subunit AB1; Ndufb4, NADH ubiquinone oxidoreductase subunit B4; Ndufb9, NADH ubiquinone oxidoreductase subunit B9; Ndufc1, NADH ubiquinone oxidoreductase subunit C1; Ndufs1, NADH ubiquinone oxidoreductase core subunit S1; Ndufs3, NADH ubiquinone oxidoreductase core subunit S3; Ndufs4, NADH ubiquinone oxidoreductase subunit S4; Ndufs6, NADH ubiquinone oxidoreductase subunit S6; Ndufs8, NADH ubiquinone oxidoreductase core subunit S8; TOM, translocase of the mitochondrial outer membrane; TOM40, translocase of outer mitochondrial membrane 40.
It has been suggested that miR-762 translocates into mitochondria in murine cardiomyocytes upon anoxia/reoxygenation treatment and decreases complex I activity and intracellular ATP levels, increases ROS levels, and enhances apoptotic cell death through suppressing the expression of NADH ubiquinone oxidoreductase chain 2 (ND2), which is a subunit of complex I [82]. Intriguingly, the administration of an inhibitor (antagomir) of miR-762 ameliorated myocardial ischemia/reperfusion injury in mice. Another example of the negative regulation of ETC complexes is provided by miR-181c, which binds to the 3′ UTR of cytochrome c oxidase subunit 1 (COX1) mRNA, encoding a subunit of complex IV and suppressing COX1 translation in rat cardiac myocytes. The overexpression of miR-181c dramatically decreased COX1 protein levels, leading to complex IV remodeling and altering mitochondrial function, including by precipitating augmented ROS production [17]. Notably, the systemic administration of miR-181c in rats diminished exercise capacity and caused heart failure [101]. For patients with heart failure with a preserved ejection fraction (HFpEF), exercise training is a treatment that commonly used to improve such patients' aerobic capacity and quality of life. A recent study has shown that the circulating level of miR-181c is significantly higher in low responders to exercise training, suggesting  Table 2 in mitochondrial ETC and OXPHOS are illustrated herein. miRNAs, tRFs, lncRNAs, and circRNAs are represented in blue, red, green, and orange, respectively. Ndufa1, NADH ubiquinone oxidoreductase subunit A1; Ndufa2, NADH ubiquinone oxidoreductase subunit A2; Ndufa3, NADH ubiquinone oxidoreductase subunit A3; Ndufab1, NADH ubiquinone oxidoreductase subunit AB1; Ndufb4, NADH ubiquinone oxidoreductase subunit B4; Ndufb9, NADH ubiquinone oxidoreductase subunit B9; Ndufc1, NADH ubiquinone oxidoreductase subunit C1; Ndufs1, NADH ubiquinone oxidoreductase core subunit S1; Ndufs3, NADH ubiquinone oxidoreductase core subunit S3; Ndufs4, NADH ubiquinone oxidoreductase subunit S4; Ndufs6, NADH ubiquinone oxidoreductase subunit S6; Ndufs8, NADH ubiquinone oxidoreductase core subunit S8; TOM, translocase of the mitochondrial outer membrane; TOM40, translocase of outer mitochondrial membrane 40.
MitomiRs also negatively regulate ATP production by targeting complex V subunits. In a mouse model of type I diabetes mellitus, it has been shown that miR-378a redistributes to interfibrillar mitochondria (IFM) and downregulates the expression of ATP synthase F0 subunit 6 (ATP6), which encodes a subunit of complex V. The administration of an inhibitor of miR-378a alleviated ATP6 expression in IFM and heart failure in this mouse model [83]. In addition, miR-378a levels increased in subsarcolemmal mitochondria (SSM) but not in IFM, and miR-378a was responsible for modulating ATP6 expression in a mouse model of type II diabetes mellitus (db/db mice) [84]. Intriguingly, the knockout (KO) of miR-378a rescued complex V activity and cardiac function in db/db mice, thus suggesting that miR-378a may be a potential therapeutic target for treating diabetes mellitus. Further study on the molecular mechanism by which miR-378a targets mitochondria will be important for its clinical application [97].
For the positive regulation of ETC complexes, miR-21, a mitomiR, interacts with the open reading frame of the mRNA of cytochrome b (CYTB), encoding a subunit of complex III, and promotes the translation of CYTB to reduce ROS production. Experiments using spontaneous hypertensive rats have suggested that this mitomiR reduces blood pressure and suppresses cardiac hypertrophy [85]. MiR-1 is another example of a mitomiR that enhances the mitochondrial translation of ETC complexes. The muscle-specific miR-1, which is induced during myogenesis, directly and specifically recruits mRNAs encoding NADH ubiquinone oxidoreductase core subunit 1 (ND1), a subunit of complex I, and COX1 to mitochondrial ribosomes and promotes the translation of these mRNAs. This function of miR-1 is dependent on AGO2 but is independent of an AGO2 partner glycine-tryptophan protein of 182 kDa (GW182) [86]. Another mitomiR, miR-5787, promotes the translation of cytochrome c oxidase subunit 3 (COX3) in tongue squamous cell carcinoma (TSCC) cells. In Cal27 and Scc25 TSCC cells, the decrease in miR-5787 expression contributes to the shifting of glucose metabolism from OXPHOS to glycolysis and enhances cisplatin resistance [87].
It has also been demonstrated that several mitomiRs affect ETC complexes via regulating mitochondrial gene expression through a route that does not occur post-transcription. The mitomiR miR-2392 has been reported to directly regulate mtDNA transcription. miR-2392 recognizes target sequences in the H-strand and partially represses polycistronic mtDNA transcription in a cell-type-specific fashion. As a consequence, miR-2392 represses the expression of ETC complex subunits, such as ND4, COX1, and cytochrome c oxidase subunit 2 (COX2), to enhance anaerobic respiration in TSCC cells [88].
Mitochondria-associated miRNAs that are located in the cytoplasm modulate mitochondrial functions by targeting nuclear genome-encoded mitochondrial transcripts in the cytoplasm (Table 2 and Figure 3). Some studies have reported that miRNAs affect ATP synthesis by altering the expression of the ETC complex and complex V subunits. For instance, one brain-specific miRNA, miR-338, lowers the expression of the nuclear genome-encoded cytochrome c oxidase subunit 4 (COX4), a subunit of complex IV, after its binding to the 3 -UTR of its mRNA [89]. When an miR-338 inhibitor was transfected into axons of neural cells, metabolic oxygen consumption was increased by about 50% when compared with nontargeting cells. Other examples include miR-210, which modulates mitochondrial functions during hypoxia by directly reducing the expression levels of cytochrome c oxidase assembly factor heme A:farnesyltransferase COX10, which facilitates the assembly of complex IV, and miR-101, which inhibits the replication of herpes simplex virus-1 (HSV-1) by downregulating the expression of ATP synthase F1 subunit β (ATP5B), a subunit of complex V [90,91]. miR-663 has been shown to increase the expression levels of ETC complex (I, II, III, and IV) subunits and assembly factors, thereby stabilizing the ETC supercomplexes. Authors have proposed that miR-663 may exert tumor-suppressive effects in breast cancer through enhancing ETC and OXPHOS activity [92]. Although ubiquinol cytochrome c reductase complex assembly factor 2 (UQCC2) is a potential target of miR-663, the mechanism by which miR-663 controls the expression of a wide range of ETC-related proteins remains unclear.
Interestingly, recent studies show the involvement of miRNAs in maladaptive retrograde signaling in response to OXPHOS dysfunction caused by pathogenic mtDNA mutations [103][104][105]. Dysfunction in OXPHOS caused by various pathological mutations modulates the levels of ROS-sensitive miRNAs. These miRNAs regulate the expression of mitochondrial tRNA-modifying enzymes [103,104]. By regulating the modification of mt-tRNAs, these miRNAs can potentially affect the speed and accuracy of mitochondrial translation, which can help mitigate the stoichiometric imbalance between mtDNA-and nuclear genome-encoded ETC complexes and complex V subunits originated through pathological mtDNA mutation. In addition, pathological mtDNA mutation can lead to the dysregulation of other miRNAs, which significantly affects the nuclear expression and potentially explains certain characteristics of mtDNA disease [106].

Roles of tRNA-Derived Small Fragments in the Regulation of Mitochondrial ETC Complexes
Since many mitochondrial diseases are caused by mutations in mt-tRNA genes, it follows that mitochondrial translation is involved in the pathophysiology of cells and tissues. Human mtDNA is a circular and double-stranded DNA consisting of 16,569 base pairs that encodes 37 genes. Thirteen of these genes are translated through specific decoding via mitochondrial ribosomes to the essential subunits of mitochondrial ETC complexes I, III, IV, and also complex V [107]. Mutations in mitochondrial DNA, particularly in mt-tRNA, are associated with syndromes such as myoclonic red fiber epilepsy (MERRF) and cardiomyopathy [108].
As tRNA-derived small fragments (tRFs) have recently been discovered to be regulators of mitochondrial biology, there are only a few reports on the roles of tRFs in relation to ETC complex regulation (Table 2 and Figure 3). As previously mentioned, mtDNA mutations causing OXPHOS dysfunction lead to changes in the levels of certain miRNAs controlling the modification of mt-tRNAs. One well-known example is the m.3243A>G mutation in the mitochondrially encoded tRNA leucine 1 (UUA/G) (MT-TL1) gene that encodes mt-tRNA Leu(UUR) . It is one of the most common mitochondrial pathogenic mutations, with a carrier frequency estimated in the range between 0.95 and 18.4 per 100,000 people in northern European populations [109][110][111]. This mutation is associated with several diseases such as MELAS, chronic progressive external ophthalmoplegia (CPEO), diabetes mellitus, and deafness. Although the manifestation of diseases varies significantly in different tissues based on the proportion of mutant mtDNA molecules present, patients with an m.3243A>G mutation in the MT-TL1 gene often show a severe, pre-existing deficiency in ETC complexes I and IV [112,113]. Accumulated data suggest that a deficiency in the aminoacylation of mutant mt-tRNA Leu(UUR) and a lack of taurine-containing modifications of its anticodon wobble position affecting the recognition of UUG codons (U34) may constitute the origin of a mitochondrial translation defect [114][115][116][117][118]. This leads to a decrease in steady-state levels of ETC complexes and an affected mitochondrial respiration rate. In addition, ROS-sensitive miRNAs, termed miR-9/9*, are induced by ETC complex deficiency, which downregulates the expression of U34-modification enzymes and leads to the hypomodification of several mt-tRNA species, including mt-tRNA Glu(UUC) . This causes the accumulation of mt-i-tRF Glu(UUC) , which is produced by the action of the nuclease DICER. An increase in mt-i-tRF Glu(UUC) downregulates the expression of the nuclear gene mitochondrial pyruvate carrier 1 (MPC1), promoting the increase in extracellular lactate levels and leading to lactic acidosis, which is one of the well-known phenotypes of MELAS [57]. This report was the first to demonstrate the involvement of tRF in the phenotype of ETC complex deficiency by clarifying the mitochondria-nucleus crosstalk controlled by small noncoding RNAs, while the hypomodification at important sites in mature tRNAs is assumed to be the underlying mechanism of the clinical manifestation of this disease with ETC complex deficiencies [119][120][121][122]. Furthermore, a recent study reported that 5 -tRNA halves derived from tRNA His(GTG) , tRNA Glu(CTC) , and tRNA Glu(TTC) (tiRNA-5 His(GTG) , tiRNA-5 Glu(CTC) , and tiRNA-5 Glu(TTC) , respectively) modulate MPC1 expression in neonatal rat pancreatic β cells and that an inhibitor cocktail of these tRNA halves suppresses the mitochondrial respiration, proliferation, and insulin secretion of these cells, suggesting their importance in postnatal β cell maturation. tiRNA-5 His(GTG) and tiRNA-5 Glu(CTC) are assumed to mediate these functions through mitochondria since they were found in a mitochondrial fraction [93].

Roles of Long Noncoding RNAs in the Regulation of Mitochondrial ETC Complexes
mt-lncRNAs, including both RNAs derived from mtDNA and nuclear genome-encoded lncRNAs transported into mitochondria, collaborate with transcription factors and other epigenetic regulators to regulate mitochondrial gene expression and function. These mt-lncRNAs play a crucial role in multiple gene-regulatory networks, serving as possible epigenetic mediators that coordinate nuclear and mitochondrial functions [62,123] ( Table 2 and Figure 3). One mt-lncRNA that is reported to regulate the expression of ETC complexes is metastasis-associated lung adenocarcinoma transcript 1 (MALAT1). MALAT1 is a nuclear genome-encoded lncRNA. A recent study indicated that MALAT1 is located in mitochondria in HepG2 hepatocellular carcinoma (HCC) cells. RNA reverse transcription-associated trap sequencing (RAT-seq) and chromatin isolation via an RNA purification (ChIRP) assay showed that MALAT1 interacts with multiple sites on mtDNA, including the D-loop region and the COX2 and NADH ubiquinone oxidoreductase core subunit 3 (ND3) gene loci. MALAT1 modulates the CpG methylation of mtDNA and the expression of mtDNA-encoded ETC-related genes, such as COX1, COX2, ND1, and ND3 genes. MALAT1 depletion impairs mitochondrial biogenesis and intracellular ATP synthesis; suppresses HCC cell proliferation, migration, and invasion; and induces mitochondria-associated apoptosis. Thus, MALAT1 may be an important therapeutic target for HCC, and the significance of MALAT1's function in mitochondria requires further study [18].
Another nuclear-genome-encoded lncRNA, survival-associated mitochondrial melanomaspecific oncogenic non-coding RNA (SAMMSON), has been shown to localize in mitochondria in melanoma cells. In these cells, SAMMSON interacts with p32, a master regulator of mitochondrial homeostasis and metabolism, and facilitates the mitochondrial localization of p32. The silencing of SAMMSON decreases the expression levels of mtDNA-encoded ETC complex subunits and the activities of ETC complexes I and IV and suppresses tumorigenesis via enhancing apoptosis [94,95]. Meanwhile, a recent study showed that SAMMSON silencing increases the expression levels of ETC complexes, cellular respiration, and mitochondrial replication in doxorubicin-resistant MCF-7 cells [96]. This finding suggests context-dependent functions for SAMMSON, although further analysis is required to clarify the binding partners and molecular mechanism of this lncRNA.
In addition, it has been indicated that some mitochondria-associated lncRNAs function as miRNA sponges in order to modulate the expression of ETC complexes. For example, it has been suggested that the lncRNA potassium voltage-gated channel subfamily Q member 1 overlapping transcript 1 (Kcnq1ot1) acts as a sponge for miR-378a and upregulates ATP6 expression and complex V activity in cardiomyocytes [97]. Another lncRNA, cytoplasmic endogenous regulator of oxidative phosphorylation 1 (Cerox1), binds to and inhibits miR-488-3p to enhance the expression of multiple ETC complex I subunits and mitochondrial respiration in N2A neuroblastoma cells [98].

Roles of Circular RNAs in the Regulation of Mitochondrial ETC Complexes
Recent studies have indicated that circRNAs can act as scaffolds in protein complex assembly, and some circRNAs regulate mitochondrial functions through direct binding to subunits of ETC complexes and complex V (Table 2 and Figure 3). For instance, the mitochondria-located circRNA circular RNA pumilio-1 (circPUM1) has been reported to act as a scaffold in order to regulate the spatial conformation of ETC complex III in KYSE30 and KYSE410 esophageal squamous cell carcinoma (ESCC) cells [99]. circPUM1 binds to ubiquinol cytochrome c reductase core protein 2 (UQCRC2) and modulates the formation of the UQCRC1-UQCRC2 dimer in mitochondrial complex III, which helps pump hydrogen from the mitochondrial matrix to the mitochondrial intermembrane space [124]. The silencing of circPUM1 leads to an intense interaction between UQCRC1 and UQCRC2, whereas circPUM1 overexpression disrupts it. Furthermore, circPUM1 silencing decreases ATP synthesis, whereas circPUM1 overexpression promotes it. Conversely, ROS levels are increased through circPUM1 silencing and decreased via circPUM1 overexpression. These findings suggest that circPUM1 plays a critical role in the proper assembly of the UQCRC1-UQCRC2 dimer and, therefore, OXPHOS. Intriguingly, circPUM1 silencing induces pyroptosis, a kind of programmed cell death, suggesting that this circRNA is a potential target of ESCC treatment. Furthermore, it has been reported that a circRNA regulates mitochondrial ROS output through interacting with a complex V subunit, namely, ATP5B [76]. Steatohepatitis-associated circRNA ATP5B regulator (SCAR) is a mitochondrial genome-encoded circRNA that is downregulated in fibroblasts of liver tissues from patients with non-alcoholic steatohepatitis (NASH) cirrhosis. SCAR binds to ATP5B, which acts as a regulator of the mitochondrial permeability transition pore (mPTP) complex. The SCAR-ATP5B complex blocks mPTP from cyclophilin D, a facilitator of mPTP opening, and shuts down mPTP, thereby inhibiting mitochondrial ROS output. Interestingly, circRNA SCAR alleviates high fat diet-induced cirrhosis in mice, suggesting that the manipulation of circRNA SCAR expression may be applicable to therapeutic strategies of NASH.
Moreover, mitochondrial circRNA for translocating phosphoglycerate kinase 1 (mcPGK1) has been indicated to be involved in metabolism reprogramming from OXPHOS to glycolysis. mcPGK1 promotes the mitochondrial import of a key glycolytic enzyme, PGK1, by facilitating the interaction between PGK1 and the translocase of outer mitochondrial membrane 40 (TOM40), which forms the central channel of the translocase of the mitochondrial outer membrane (TOM complex). mcPGK1 is highly expressed in primary liver tumor-initiating cells (TICs) and is involved in TIC self-renewal, indicating the significance of mcPGK1 in clinical management of liver cancer [100].

Conclusions
In the present review, we have described multiple classes of noncoding RNAs and their regulatory pathways involved in the mitochondrial electron transport chain and OXHPOS (Table 2 and Figure 3). Clarifying the mechanisms of the noncoding RNAs regulating the mitochondrial electron transport chain and OXHPOS will help identify therapeutic targets for various diseases such as MERRF, MELAS, and cardiomyopathy in which OXPHOS plays a crucial role. The identification of new noncoding RNAs affecting the mitochondrial electron transport chain, OXPHOS, and their related molecules will allow us to further understand pathophysiology of the diseases. Although some noncoding RNAs mentioned in this review can be targets for innovative treatment or prophylactic agents for OXPHOSrelated conditions, they are still far from clinical application. Both basic and clinical research on noncoding RNAs and the pathways by which they regulate the mitochondrial electron transport chain and OXPHOS would be required for the discovery of novel therapeutic approaches for OXPHOS-related diseases. Funding: This work was partially supported by grants from the Japan Society for the Promotion of Science, Japan [23K08837 (to T.T.); 21H02981 and 22K19572 (to K.I.); and 21H04829 (to S.I.)] and by the Takeda Science Foundation (to K.I. and S.I.).